Currently there exists no vaccine for the prevention of infection with the ubiquitous gastric pathogen Helicobacter pylori, and drug therapy for the infection is complicated by poor patient compliance, the high cost of treatment, and ineffectiveness against drug resistant strains. Clearly a new medical advancement is required to reduce the incidence of peptic ulcer disease and stomach cancer, two conditions caused by infection with H. pylori. Pre-clinical studies have suggested that vaccination could not only prevent infection, but could also be used to resolve active infections. An effective vaccine may also protect against re-infection as well as be an impediment to transmission in at-risk populations. Antex Biologics is an industry leader in H. pylori vaccine development, and is committed to bringing an H. pylori vaccine to the market. The company has performed two Phase 1 clinical trials with a formalin-inactivated Helicobacter pylori Whole Cell (HWC) vaccine, which was given orally in combination with the adjuvant mLT(R192G), a mutant of E. coli heat-labile toxin. Pre-clinical results suggest that the use of an adjuvant with HWC is required for vaccine efficacy. In an effort to bring to market an optimally safe, stable, and potent vaccine, we propose in this application to evaluate clinically two new aspects of the vaccine formulation, namely 1) a lyophilized rather than liquid active ingredient (HWC), and 2) a novel double mutant form of LT (dmLT) that has reduced enterotoxic activity compared to the single mutant mLT(R192G), yet retains full adjuvant activity. The lyophilization process for HWC will be selected based upon in vitro testing of the product for characteristics such as moisture content and reconstitution behavior, and in vivo based upon animal immunization and challenge experiments. The final formulation selected will be used to advance understanding of the vaccine mechanism of action through the use of CD4, antibody, and IFN-gamma knockout mouse studies, and flow cytometric studies examining the nature of the gastric T cell infiltrate in vaccinated and challenged mice. cGMP lots of lyophilized HWC and dmLT will be produced and a Phase 1 dose escalation safety trial in uninfected and infected, asymptomatic subjects will be performed. Because dmLT has yet to be evaluated clinically, the maximum tolerable dose of dmLT alone will be determined first in both populations. Following dmLT dose selection, the maximum tolerable dose of lyophilized and reconstituted HWC in the context of a fixed dose of dmLT will then be determined, again in both populations. Diarrhea is the most likely adverse event that will define tolerance limits. A panel of immunological tests will be performed on subjects receiving the full vaccination series with HWC + dmLT, including serum and mucosal antibodies to HWC and dmLT, and PBMC proliferation and cytokine production in response to restimulation with H. pylori lysate. Diagnostic testing will also be performed on infected subjects post-vaccination to assess changes in their colonization state. This trial will position the vaccine for more highly powered Phase 1 or early Phase 2 trials, and thus will move H. pylori vaccination one step closer to being a reality.